As is known, once forming stem-loops, the oligonucleotides will be more stable than the single-stranded ones. And mRNA with stem-loop at its 3' or 5' end often get a longer lifetime than the linear one owe to the stem-loop's resistance to exonuclease. Our team tend to design a series of stem-loops each followed by the same endonuclease site and are transcribed as one polycistron. Once digested by endonuclease and seperate into several independent fragments, cistrons with different free energy stem-loops will get different stability, thus influence the amount of expressed proteins. In this way, we can decouple the expression level of upstream and downstream genes of the same operon by simply designing different stem-loops. Futhermore, with quantitative free energy of stem-loops, we even can achieve the ratio expression of target proteins. It is a creative regulating method and we attempt to provide a series of standard regulation parts for others.

We have developed our own software which provides a database of stem loops. When users upload their own sequence, the software can find out corresponding stem loops and store them into the database. Moreover, when users need proteins expressed in specific quantity, such as inputting the ratio of two different proteins, the software can provide suitable stem loops according to the ratio. The relationship between the quantitative expression of protein and the folding energy is based on our model.